Proteins are the essential building blocks of living cells. They play a central role in medicine and pharmacy – but important aspects of their behaviour have not yet been fully explained. Physicists at TU Wien (Vienna) have now proposed an answer to an important question in molecular biology: How is it possible that proteins sometimes distribute themselves homogeneously in a liquid, but sometimes form self-contained droplets without mixing with the surrounding liquid? The decisive factor here is the distribution of electrical charges.

When liquids suddenly separate into two parts

“Our starting point was an observation often been made in biology,” says Emanuela Bianchi from the Institute of Theoretical Physics at TU Wien. “Liquids that contain proteins sometimes split spontaneously into droplets with an extremely high protein concentration and a remaining liquid that hardly contains any proteins at all. This is referred to as two different liquid phases that do not mix.”

This can be a problem in medicine. “When delivering a drug, for example, you actually want the active molecules to bind with the target proteins in the patient body and not bind together forming small droplets,” says Daniele Notarmuzi (TU Wien). Whether a particular protein tends to form droplets or mixes well with the surrounding liquid depends on various factors: For example, on the concentration of the protein particles, the pH value of the liquid and the temperature. So far, however, it has only been possible to rely on empirical reports and there has been no systematic explanation of the phenomenon.

The team at TU Wien, however, has a lot of experience in calculating the complex interaction of many small particles. “We know that electrical charge can play an important role in the collective behaviour of small particles,” says Emanuela Bianchi. “So we investigated whether the behaviour of proteins can be explained this way.”

Proteins as little spheres with an electrical charge

Emanuela Bianchi and Daniele Notarmuzi viewed the proteins mathematically as small spheres that can be either electrically neutral or electrically charged. The charge can be distributed in different ways: “For example, they may be spheres with a largely negatively charged surface, in which only small regions at the north and south pole is positively charged,” says Notarmuzi. “Spheres with such a charge distribution will arrange themselves in a different pattern than, for example, spheres that have larger positively charged regions on their surface.”

When such spherical proteins come into contact with each other, a positively charged north or south pole, for example, will preferentially attach to the negatively charged equator of a neighbouring particle, because opposite charges attract each other. This in turn determines how a third particle can attach itself and how stable this configuration is – and so on, until a large protein cluster of many particles has formed.

Details of the charge distribution in the proteins can therefore have a major impact on which structures and patterns emerge, when many proteins interact. This also determines whether these structures form stable droplets, or whether they break up immediately and dissolve in the surrounding liquid.

Physics makes biology predictable

The computational model from TU Wien combines two different areas of physics: on the one hand, electrostatics, which can be used to precisely calculate the forces between charged particles, and on the other hand, statistical physics, which provides information about the collective behaviour of the particles that these forces can lead to.

“We have successfully used this approach to explain under which circumstances the proteins are mixed and under which circumstances they form droplets,” says Emanuela Bianchi. For example, it can sometimes be sufficient to change the pH value of the surrounding liquid in order to switch the behaviour of the proteins. It is also possible for experimental biologists to modify the molecular blueprint of the proteins: “Different amino acids lead to different distributions of electrical charge. It is possible to genetically modify in vitro proteins to obtain a mutated version that then exhibits better mixing behaviour,” says Daniele Notarmuzi.

“Our aim was to develop a solid physical foundation on which we can now pursue questions that are important for other sciences,” says Emanuela Bianchi. “We were able to show that electrical charge plays the decisive role. We hope that, in many cases, biology and pharmacy will no longer have to rely on trial and error but will be able to determine what the best solution is based on our understanding of the physical principles governing this phenomenon.”

Original publication

D. Notarmuzi and E. Bianchi: Liquid-liquid phase separation driven by charge heterogeneity, Communications physics, 412 (2024)., opens an external URL in a new window